The analysis of a substance to determine its composition may be necessary for many applications, including toxicology, forensics and environmental testing, as well as food and drug research. Often, samples to be analyzed are analyzed for the presence of numerous different analytes of interest. Such samples may, for example, be in the form of bodily fluids taken from test subjects, which fluids often include both drug metabolites of interest, as well as irrelevant endogenous ions from the test subject. Correctly determining the presence or absence of a large number of analytes of interest from complex substances can be difficult and time-consuming.
Mass spectrometers are often used for producing a mass spectrum of a sample to find its composition. For example, chromatographic equipment such as a liquid chromatograph is frequently used to elute ions from a sample into the mass spectrometer over a period of time and recursive MS techniques can be utilized to identify the structure of various analytes contained within portions of the eluted sample.
In most instruments with MS/MS capabilities, the process of generating a mass spectrum, selecting a precursor ion and generating an MS/MS spectrum of product ions can be performed in an automated mode. Current automated protocols in tandem mass spectrometry generally perform a single MS of a sample (e.g., a survey scan) to identify candidates within the sample for further processing, and select among all species observed a subset of precursor ions for analysis by MS/MS. This is normally achieved by ionizing the sample and separating ions of differing masses and recording their relative abundance by measuring intensities of ion flux in a survey scan. For example, with time-of-flight mass spectrometers, ions are pulsed to travel a predetermined flight path. The ions are then subsequently recorded by a detector. The amount of time that the ions take to reach the detector, the “time-of-flight,” may be used to calculate the ion's mass to charge ratio, m/z. Additional information can then be obtained by fragmenting the selected precursor ion(s) via CID (collision induced dissociation) in a collision cell (or other means) to generate an MS/MS spectrum.
In selecting the precursor ions for further processing, current automated protocols generally focus on the most abundant precursor ions within the eluted sample, as determined by their detected intensity in the survey scan so as to maximize the number of high quality, identifiable MS/MS spectra. That is, both the precursor ions selected for further processing and the parameters used during MS/MS are generally selected to maximize the chances of being able to identify the structure of various precursor ions (and their constituents). By way of example, the most-intense 20 precursor ions detected via the survey scan are ranked based on their intensity and serially processed from most-intense to least-intense in one or more downstream product ion stages. Examples of this mode of acquisition include Information Dependent Acquisition (IDA) or Data Dependent Acquisition (DDA).